Previously, cDNAs were constructed to encode various truncated versions of the genome RNA of respiratory syncytial virus (RSV) in which the genome termini and putative transcription signals were retained but the viral genes were deleted and replaced with a reporter gene such as that for bacterial chloramphenicol acetyl transferase (CAT). When introduced into RSV-infected cells, the prototype RSV-CAT minigenome was replicated, transcribed and packaged into virus-like particles. This represents the first type of system in which genome-like RNAs of a nonsegmented negative strand RNA virus could be introduced into the viral replication cycle and rendered biologically active. Here, this system was used to identify and characterize the cis-acting replication and transcription signals in the RSV genome and to study the mechanism of gene expression. It also was adapted for the analysis of protein function and for the generation of infectious RSV from a cDNA of the complete genome (accompanying reports).